ABSTRACT Potassium channels are critical for life, playing a variety of physiological roles including control of solute balance, cell volume, and cellular excitability. In excitable cells potassium channels are involved in setting membrane potential and modulation of action potentials. Slack channels, encoded by KCNT1, are members of the Slo family of potassium channels, comprised of Slo1 (Maxi-K), Slo2.1 (Slick), Slo2.2 (Slack) and Slo3, a sperm-specific channel. Recently, multiple studies have identified ~30 mutations in Slack channels that are associated with a variety of epilepsies, most frequently Malignant Migrating Partial Seizures of Infancy (MMPSI). MMPSI are extremely severe, pharmacoresistant, and typically fatal during the first year of life. Slack variants associated with these epilepsies are found in three clusters: the pore of the channel, and in two separate carboxy-terminal regulatory regions, Regulator of Conductance of K+ (RCK) and NAD+ binding domains. Interestingly, these mutations are dominant, gain-of-function mutations resulting in a higher degree of Slack channel activity. Although the exact mechanism by which over-activity of Slack leads to MMPSI and other, less severe epilepsies is not understood, we hypothesize that specific inhibitors of Slack channels may provide a viable anti-epileptic strategy. Thus, the aims of this proposal are to build a hit-discovery and characterization infrastructure focused on providing an ability to conduct an 110,000 compound screen on wild-type Slack and 3 MMPSI-associated mutants, one from each of the three clusters (R21 Phase). We will utilize this infrastructure and a focused hit-to- leads medicinal-chemistry effort, including assays and strategies geared toward enhancing drug-likeness, to support the discovery and characterization of Slack-specific inhibitors (R33 Phase). The goal of the proposed research is to discover and validate at least two small molecule scaffolds, providing a platform for development of in vivo-active Slack inhibitors and a foundation for a therapeutically focused lead optimization effort.